Detection of hepatitis E virus RNA in rats caught in pig farms from Northern Italy

Hepatitis E virus (HEV) strains belonging to the Orthohepevirus genus are divided into four species (A–D). HEV strains included in the Orthohepevirus A species infect humans and several other mammals. Among them, the HEV‐3 and HEV‐4 genotypes are zoonotic and infect both humans and animals, of which, pigs and wild boar are the main reservoirs. Viruses belonging to the Orthohepevirus C species (HEV‐C) have been considered to infect rats of different species and carnivores. Recently, two studies reported the detection of HEV‐C1 (rat HEV) RNA in immunocompromised and immunocompetent patients, suggesting a possible transmission of rat HEV to humans. The role of rats and mice as reservoir of HEV and the potential zoonotic transmission is still poorly known and deserves further investigation. To this purpose, in this study, the presence of HEV RNA was investigated in the intestinal contents and liver samples from 47 Black rats (Rattus rattus) and 21 House mice (Mus musculus) captured in four pig farms in Northern Italy. The presence of both Orthohepevirus A and C was investigated by the real‐rime RT‐PCR specific for HEV‐1 to HEV‐4 genotypes of Orthohepevirus A species and by a broad spectrum hemi‐nested RT‐PCR capable of detecting different HEV species including rat HEV. The intestinal content from two Black rats resulted positive for HEV‐C1 RNA and for HEV‐3 RNA, respectively. None of the House mice was HEV RNA positive. Sequence analyses confirmed the detection of HEV‐C1, genotype G1 and HEV‐3 subtype e. The viral strain HEV‐3e detected in the rat was identical to swine HEV strains detected in the same farm. Liver samples were negative for the detection of either rat HEV or HEV‐3.     

Hepatitis E virus in Common voles (Microtus arvalis) from an urban environment,Hungary: Discovery of a Cricetidae‐specific genotype of Orthohepevirus C

Hepatitis E virus is a major causative agent of acute hepatitis worldwide. Despite its zoonotic potential, there is limited information about the natural chain of hepevirus infection in wildlife, and the potential reservoir species. In this study, we performed a HEV survey by heminested RT‐PCR on rodent samples from an urban environment (in the city of Pécs, Hungary) and investigated the prevalence of the virus among these native rodent species (Apodemus agrarius, Apodemus flavicollis, Apodemus sylvaticus, Microtus arvalis and Myodes glareolus). HEV was detected exclusively in Common voles (M. arvalis), in 10.2% of screened voles, and 3.2% of all investigated samples from all species. Based on the phylogenetic analysis, our strain showed the closest homology with European Orthohepevirus C strains detected previously in faecal samples of birds of prey and Red fox, supporting the possibility of the dietary origin of these strains. In addition, our samples showed close phylogenetic relation with a South American strain detected in Necromys lasiurus (Cricetidae), but separated clearly from other Muridae‐associated strains, suggesting the presence of a Cricetidae‐specific genotype in Europe and South‐America. Based on these results, we hypothesize the reservoir role of M. arvalis rodents for the European Cricetidae‐specific Orthohepevirus C genotype.     

Hepatitis E virus in wild rabbits and European brown hares in Germany

Recently, a change of hepatitis E from being a typical travel‐associated disease to an autochthonous zoonosis in Germany was observed. An increasing number of autochthonous infections with the hepatitis E Virus (HEV) have been recognized in developed countries. Venison from wild boar is already known to be a potential source of infection, if not prepared properly by the consumer. In Germany, certain wild animals are known to be a reservoir for HEV. However, current information is missing about European brown hares (Lepus europaeus) and wild rabbits (Oryctolagus cuniculus). Thus, a total of 833 hunting‐harvested animals (European brown hares n = 669; wild rabbits n = 164) were tested for the occurrence of HEV RNA and HEV antibodies. For this, liver and blood specimens were taken after hunts in six German federal states. HEV antibodies were found by ELISA in 2.2% (624/14) of European brown hares, but no HEV RNA was detectable by nested real‐time RT‐PCR. In contrast, a seroprevalence of 37.3% (126/47) was observed for wild rabbits, and 17.1% (164/28) of the samples were HEV RNA positive. Genomic analysis revealed that these partial sequences clustered within the rabbit clade of HEV‐3 genotype. In addition, one rabbit sequence segregated into subtype 3g of HEV‐3. Highest seroprevalences for hares and rabbits were detected in the federal states of Bavaria and of Schleswig‐Holstein, respectively. Comparing urban, rural and insular areas, the highest seroprevalence was shown for wild rabbits in rural areas and for European brown hares on the northern island Fehmarn. This study provides evidence that European brown hares and wild rabbits from Germany can be infected with HEV. The different prevalences indicate that wild rabbits are a potential reservoir for HEV in Germany, whereas European brown hares seem to be only of minor importance for the epidemiology of HEV.     

Beyond abundance: How microenvironmental features and weather influence Bartonella tribocorum infection in wild Norway rats (Rattus norvegicus)

Norway rats (Rattus norvegicus) inhabit cities worldwide and carry a number of zoonotic pathogens. Although many studies have investigated rat‐level risk factors, there is limited research on the effects of weather and environment on zoonotic pathogen transmission ecology in rats. The objective of this study was to use a disease ecology approach to understand how abiotic (weather and urban microenvironmental features) and biotic (relative rat population abundance) factors affect Bartonella tribocorum prevalence in urban Norway rats from Vancouver, British Columbia, Canada. This potentially zoonotic pathogen is primarily transmitted by fleas and is common among rodents, including rats, around the world. During a systematic rat trap and removal study, city blocks were evaluated for 48 environmental variables related to waste, land/alley use and property condition, and rat abundance. We constructed 32 weather (temperature and precipitation) variables with time lags prior to the date we captured each rat. We fitted multivariable logistic regression models with rat pathogen status as the outcome. The odds of a rat testing positive for B. tribocorum were significantly lower for rats in city blocks with one or more low‐rise apartment buildings compared to blocks with none (OR = 0.20; 95% CI: 0.04–0.80; p = .02). The reason for this association may be related to unmeasured factors that influence pathogen transmission and maintenance, as well as flea vector survival. Bartonella tribocorum infection in rats was positively associated with high minimum temperatures for several time periods prior to rat capture. This finding suggests that a baseline minimum temperature may be necessary for flea vector survival and B. tribocorum transmission among rats. There was no significant association with rat abundance, suggesting a lack of density‐dependent pathogen transmission. This study is an important first step to understanding how environment and weather impacts rat infections including zoonotic pathogen ecology in urban ecosystems.     

Seroepidemiology and genotyping of hepatitis E virus in Singapore reveal rise in number of cases and similarity of human strains to those detected in pig livers

Hepatitis E virus (HEV) causes 20 million infections worldwide yearly, of which only about 3.3 million are symptomatic. In developed Asian countries, HEV strains detected in human sera and in food sources were genetically similar, suggesting that indigenous HEV infections may be largely food‐borne. To assess the burden of hepatitis E in Singapore, we performed a seroepidemiologic study of the infection. Additionally, we carried out HEV genotyping on archived, residual HEV IgM‐positive serum samples collected between 2014 and 2016 (n = 449), and on pig liver samples (n = 36) purchased from wet markets and supermarkets. Our study shows a rise in hepatitis E incidence (IgM) from 1.7 to 4.1 cases per 100,000 resident population from 2012 to 2016 and an increase in hepatitis E IgG positivity rate among residents from 14% in 2007 to 35% in 2016. Other findings also suggest the epidemiology of hepatitis E in Singapore has shifted, from it being mainly a disease imported from the Indian subcontinent, to one that is now increasingly prevalent in our resident population. Genotypes obtained from 143 human samples identified the majority to be genotype 3 (n = 121), 21 to be genotype 1 and one to be genotype 4. Further phylogenetic analyses suggest genotype 3a to be the cause of indigenous infections in residents, which showed genetic similarity to the genotype 3a strains detected in pig livers. This link between the strains in the majority of human samples and those in pig livers consumed by the public suggests a possible food‐borne route of HEV infection in Singapore.     

Epidemiology and genetic diversity of zoonotic pathogens in urban rats (Rattus spp.) from a subtropical city,Guangzhou, southern China

Commensal rats (Rattus spp.), which are globally distributed, harbour many pathogens responsible for significant human diseases. Despite this, we have a poor understanding of the epidemiology and genetic diversity of some recently neglected zoonotic pathogens, such as Leptospira spp., Bartonella spp. and hepatitis E virus (HEV), which constitute a major public health threat. Thus, we surveyed the occurrences, co‐infection and genetic diversity of these pathogens in 129 urban rats from China. For Rattus tanezumi, the prevalences of Leptospira spp., Bartonella spp. and HEV infection were 6.67%, 0% and 46.67%, respectively. The prevalences of Leptospira spp., Bartonella spp. and HEV infection were 57.89%, 9.65% and 57.89% for Rattus norvegicus respectively. Leptospira spp. and HEV infections were more likely to occur in mature R. norvegicus. Phylogenetic analyses showed that pathogenic Leptospira interrogans and Leptospira borgpetersenii might exist. We also found that Bartonella spp. showed high similarity to Bartonella elizabethae, Bartonella rochalimae and Bartonella tribocorum, which are implicated in human disease. Dual and triple infections were both detected. Moreover, dual infections with Leptospira spp. and HEV represented the most frequent co‐infection, and there was a significantly positive association between them. High genetic diversity was observed in genes segments from Leptospira, Bartonella and HEV. Our results first discover the occurrence of multiple co‐infections and genetic diversity of Leptospira, Bartonella and HEV in commensal rats from China. Altogether, the present study provides an insight into evaluating the risk of rat‐borne zoonoses in urban China.     

Zoonotic Helminths of Urban Brown Rats (Rattus norvegicus) in the UK: Neglected Public Health Considerations?

Urban brown rats (Rattus norvegicus) carry microbial human pathogens but their role as reservoir hosts for helminths of public health importance is less well known. In this study, 42 brown rats trapped on Merseyside were subject to thorough combined helminthological and pathohistological post‐mortem examination. Eggs of the rodent‐borne zoonotic nematode Calodium hepaticum were initially detected in histological sections of the livers of 9.5% of rats, but overall diagnostic sensitivity increased to 16.6% when entire liver tissue was disrupted and the resulting filtrates were examined for released eggs. In their rat host, mainly trapped inside the dockland, infections with C. hepaticum were associated with a chronic multifocal pyogranulomatous hepatitis with intralesional eggs and peripheral fibrosis. Mean intensity of hepatic C. hepaticum egg infections was 1041 eggs. This is the first report of C. hepaticum in an urban brown rat population in the UK and provides original data for liver egg burdens in this abundant commensal rodent. The zoonotic cestode Rodentolepis nana had a prevalence of infection of 14.3%. Rodent‐specific, non‐zoonotic helminths found were the spiruroid Mastophorus muris (16.0%) in the stomach, the trichuroid Trichosomoides crassicauda in the urinary bladder (31.0%); the ascarid Heterakis spumosa was the commonest helminth of the large intestine (76.2%). Many millions of brown rats inhabit cities and rural areas of the UK, and the infective stages of the zoonotic worm species, particularly C. hepaticum, are likely to be widely distributed in the environment presenting a threat to public health.     

Occupational swine exposure and Hepatitis E virus,Leptospira, Ascaris suum seropositivity and MRSA colonization in Austrian veterinarians, 2017–2018—A cross‐sectional study

We investigated the prevalence of Hepatitis E Virus (HEV), Leptospira and Ascaris suum (A. suum) seropositivity, and of nasal methicillin‐resistant Staphylococcus aureus (MRSA) colonization among Austrian practising veterinarians, and assessed the association with occupational swine livestock exposure. The 261 participants completed a questionnaire on demographics, intensity of occupational swine livestock contact and glove use during handling animals and their secretions. Participants' blood samples were tested for HEV, Leptospira and A. suum seropositivity and nasal swabs cultured for MRSA. We compared swine veterinarians (defined as >3 swine livestock visits/week) to non‐swine veterinarians (≤3 swine livestock visits/week) with regard to the outcomes through calculating prevalence ratio (PR) and 95% confidence interval (CI). Furthermore, the relationship between occupational swine livestock contact and the study outcomes was examined by age (</≥55 years) and glove usage. The prevalence of nasal MRSA colonization was 13.4% (95% CI: 9.3–17.6), of HEV seropositivity 20.8% (95% CI: 15.8–25.7) and A. suum seropositivity 44% (95% CI: 37.7–50.2). The highest anti‐leptospiral antibodies titres were 1:200 (L. hebdomadis) and 1:100 (L. autumnalis, L. caicola) found in three non‐swine veterinarians. Compared to non‐swine veterinarians, swine veterinarians were 1.9 (95% CI: 1.0–3.4) and 1.5 (95%CI: 1.0–2.3) times more likely HEV seropositive and A. suum seropositive, respectively, and 4.8 (95%CI: 2.5; 9.3) times more likely nasally colonized with MRSA. Among glove‐using veterinarians, occupational swine contact was no longer a determinant for HEV seropositivity (PR 1.6; 95% CI: 0.8–2.9). Similar was found for A. suum seropositivity, which was no longer associated with occupational swine livestock contact in the subgroup of glove using, ≥55‐year‐old veterinarians (PR: 1.07; 95% CI: 0.4–3.3). Our findings indicate that >3 occupational swine livestock visits per week is associated with HEV and A. suum seropositivity and nasal MRSA colonization and that glove use may play a putative preventive role in acquiring HEV and A. suum. Further analytical epidemiological studies have to prove the causality of these associations.     

Familial Hepatitis E Outbreak Linked to Wild Boar Meat Consumption

An HIV‐infected patient was diagnosed with acute hepatitis E infection in our hospital. An epidemiological inquiry was performed to collect demographic, food and animal exposure variables in order to identify the potential route of transmission. The patient reported that his family traditionally hunted wild boar for food. All family members were analysed for hepatitis E virus infection. Additionally, route of transmission by wild boar meat consumption and prevalence of HEV infection among wild boar from the same hunting area were investigated. In all‐family members (n = 8), HEV‐RNA was amplified. Two wild boar meat slices consumed was analysed, showing the presence of HEV. The virus isolated was consistent with genotype 3, revealing 100% homology between family members and meat. Additionally, we tested nine wild boar hunted in the same hunting area. All of them were RNA‐HEV positive, isolating the same HEV genotype 3 viral strain. We demonstrated by phylogenetic analysis zoonotic transmission of HEV by wild boar meat consumption. The prevalence of HEV infection among wild boar found in our study suggests that this species is an important route of transmission to human.     

Seroepidemiology and molecular characterization of hepatitis E virus infection in swine and occupationally exposed workers in Punjab,India

Hepatitis E virus (HEV) has two discrete epidemiological patterns: waterborne epidemics in developing countries only, caused by HEV genotype I, and sporadic zoonotic outbreaks in developing and developed countries caused by genotypes III and IV. This study was designed to investigate seroprevalence, molecular detection and the characterization of HEV by nested RT‐PCR in swine as well as the occupational risk to exposed human population in Punjab state of north‐western India. The occupational risk‐exposed group comprised of swine farmers (organized – mixed feed feeders and unorganized – swill feeders), slaughterhouse workers, sewage workers and veterinary internes. During the study period, blood and faecal samples were collected from 320 swine and 360 humans with both high and low occupational exposure risks. The overall seroprevalence of swine HEV was 65.00%, with a significantly higher seropositivity in growing pigs (2–8 months of age). The prevalence of HEV RNA in swine faecal samples by nRT‐PCR was 8.75% with a significantly higher detection in swill‐fed pigs. With humans in the high occupational exposure risk population, significantly higher anti‐HEV IgG seropositivity was observed (60.48%) as compared to control population (10.71%). Strong evidence of association between human anti‐HEV IgG seropositivity and certain occupational exposure risk groups was observed (p < 0.05). This indicates that unorganized swine farmers, slaughterhouse workers and sewage workers have higher odds of HEV infection in this study region. Percentage of nucleotide similarity between swine and human HEV isolates was less than that found in countries with zoonotic HEV outbreaks. Molecular characterization revealed the circulation of G IV and G I genotypes among swine and human population in Punjab state, respectively.     

A single‐nucleotide polymorphism in the canine cytochrome b5 reductase (CYB5R3) gene is associated with sulfonamide hypersensitivity and is overrepresented in Doberman Pinschers

Canine sulfonamide hypersensitivity (HS) has been associated with a variant in the cytochrome b₅ reductase gene (CYB5R3 729A>G), which encodes a drug‐detoxifying enzyme. Study objectives were to determine variant allele frequency in Doberman Pinschers (DOBE), a breed which may be predisposed to sulfonamide HS, and to characterize the effects of CYB5R3 729G on gene expression and function. CYB5R3 729A>G allele frequencies were compared between DOBE (n = 24) vs. non‐Doberman (non‐DOBE; n = 60) dogs. CYB5R3mRNA expression, protein expression, and reduction of sulfamethoxazole hydroxylamine were compared between banked canine liver samples of 729AA vs. GG genotype. The 729G allele was overrepresented in DOBE (1.00) vs. non‐DOBE dogs (0.567, p < .0001). mRNA and protein expressions as well as cyt b₅ reductase activity were similar between livers of AA and GG genotype. All Doberman Pinschers in this study were homozygous for CYB5R3 729G, which could contribute to this breed's apparent predisposition to sulfonamide HS. However, CYB5R3 729G does not alter sulfamethoxazole detoxification capacity, so a direct role could not be demonstrated. It is possible that this marker is linked to another contributing variant.     

Molecular Characterization and Serology of Leptospira kirschneri (Serogroup Grippotyphosa) Isolated from Urine of a Mare Post‐Abortion in Brazil

A strain of Leptospira kirschneri (serogroup Grippotyphosa) was cultured from urine of a mare post‐abortion in Brazil and characterized by serogrouping, multiple‐locus variable‐number tandem repeat analysis, PGFE, and sequencing of genes rrs and secY. Strains of L. kirschneri have apparently never been recovered from horses in tropical area, only in Europe and USA. Knowledge of local epidemiology is important to interpret genetic profiles of leptospires circulating in an area.     

Hepatitis E Virus Antibodies in Finnish Veterinarians

We investigated hepatitis E virus (HEV) infections in Finnish veterinarians engaged in different practice specialties and evaluated the effect of different background factors on HEV exposure by examining total HEV antibodies in samples collected from the participants of the 2009 National Veterinary Congress in Helsinki, Finland. Finnish veterinarians commonly have total HEV antibodies with seroprevalence of 10.2%. Of the non‐veterinarians, 5.8% were seropositive. Increasing age was associated with HEV seropositivity, and, surprisingly, the highest HEV seroprevalence (17.8%) among veterinarians was detected among small animal practitioners. Although no positive correlation between swine contacts and HEV seropositivity was found, 22.7% of veterinarians who had had needle stick by a needle that had previously been injected into a pig versus 9.0% of those who had not were seropositive, even though the finding was statistically non‐significant (P = 0.07). Our results suggest that, although contact with swine is a known risk factor for HEV infection, the sources of HEV infections are probably numerous, including travelling abroad and possibly also other reservoirs of HEV than pigs.     

Seroepidemiology of Hepatitis E in Selected Population Groups in Croatia: A Prospective Pilot Study

Hepatitis E has become an emerging infection in many European countries. We analysed the prevalence of hepatitis E virus (HEV) infection in selected population groups in Croatia. Overall HEV IgG seropositivity was 5.6%, while 1.9% participants showed IgM antibodies suggestive of recent infection. No IgM‐positive sample was positive for HEV RNA. HEV IgG antibodies were most prevalent in alcohol abusers (8.9%) and war veterans (8.6%), compared with 6.1% among injecting drug users and 2.7% in healthcare professionals. No individual with high‐risk sexual behaviour tested HEV seropositive. HEV IgG positivity increased significantly with age from 1.8% to 2.3% in individuals younger than 40 years to 11.3% in individuals older than 50 years (P = 0.023). The mean age of HEV‐positive participants was significantly higher than that of HEV‐negative participants (50.9 ± 11.8 years versus 41.2 ± 11.8 years, P = 0.008). Seroprevalence rates were significantly higher in residents of suburban and rural areas compared with residents of urban areas (14.5% versus 2.5%, P = 0.003). Additionally, an increasing prevalence of HEV IgG antibodies was observed from 1.8% in participants living in families with two household members to 12.1% in those living with more than four members (P = 0.046). Gender, marital status, educational level, sexual orientation, source of drinking water, history of blood transfusions, surgical procedures, tattooing and travelling were not associated with HEV seroprevalence. Logistic regression showed that living in suburban/rural areas was the main risk factor for HEV seropositivity (OR = 6.67; 95%CI = 1.89–25.0; AOR = 7.14, 95%CI = 1.89–25.0).     

Prevalence of Salmonella spp. in Cane Toads (Bufo marinus) from Grenada,West Indies,and their Antimicrobial Susceptibility

Cloacal swabs and caecal contents sampled from 58 cane toads (Bufo marinus) in St George’s parish, Grenada, during a 7‐month period in 2011 were examined by an enrichment and selective culture method for presence of Salmonella spp. Twenty‐four (41%) toads were positive for Salmonella spp. of which eight were Salmonella enterica serovar Javiana, and eight were S. enterica serovar Rubislaw. The other serovars were as follows: Montevideo, 6; Arechavaleta, 1; and serovar: IV:43:‐:‐, 1. The high frequency of isolation of serovar Javiana, an emerging human pathogen associated with several outbreaks in the recent years in the eastern United States, suggests a possible role for cane toads in transmission of this serovar. Although S. Rubislaw has been isolated from lizards, bats and cases of some human infections, there is no report of its carriage by cane toads, and in such high frequency. The rate of carriage of S. Montevideo, a cause for human foodborne outbreaks around the world was also over 10% in the 58 toads sampled in this study. The antimicrobial drug susceptibility tests against amoxicillin‐clavulanic acid, ampicillin, cefotaxime, ceftazidime, ciprofloxacin, enrofloxacin, gentamicin, imipenem, nalidixic acid, streptomycin, tetracycline and trimethoprim‐sulfamethoxazole showed that drug resistance is minimal and is of little concern. Antimicrobial resistance was limited to ampicillin and amoxicillin‐clavulanic acid in one isolate of S. Javiana and one isolate of S. Rubislaw. This is the first report of isolation and antimicrobial susceptibilities of various Salmonella serovars not identified previously in cane toads in Grenada, West Indies.     

Evidence of increased Hepatitis E virus exposure in Lao villagers with contact to ruminants

Although pigs are the main reservoir, ruminants have also been shown to be susceptible to hepatitis E virus (HEV). We investigated zoonotic transmission of HEV in rural settings of Lao People's Democratic Republic (Lao PDR) where humans are in close contacts with ruminants and where pigs are rare. Villagers with (n = 171, risk group) and without (n = 155, control group) cattle were recruited in seven villages in Vientiane Capital. Owners of pigs were excluded. Blood, as well as information on socio‐demographics, animal contact, dietary habits and awareness of zoonoses were collected to assess risk factors. Blood and rectal swabs were collected from cattle (n = 173) and other ruminants (27 goat, 5 buffaloes) to measure anti‐HEV antibody and virus prevalence. A similar anti‐HEV antibody seroprevalence was found in cattle (6.8%) and other ruminants (8%). HEV RNA was detected in none of the animal rectal swabs and human sera. Anti‐HEV IgG seroprevalence was higher in cattle farmers than in the control group (59.1% vs. 43.9%, p = 0.008) and increased significantly with age. Other risk factors included male gender, close contact with cattle and consumption of undercooked meat. We find that HEV is highly endemic in rural Laos and provide first evidence that HEV circulates in free‐roaming ruminants with open access to village water sources. Despite some awareness about hygiene, villagers are likely constantly exposed to zoonotic diseases by dietary and lifestyle habits. Cattle farmers had a higher risk of HEV infection than other villagers. Our study highlights the need to raise the awareness of the rural population about water‐ and food‐borne pathogens, and about the role of cattle as a possible source of infection. The knowledge gained on local risk factors and husbandry conditions should guide future awareness raising campaigns and promote appropriate hygienic measures including handwashing and the consumption of safe food and water.     

Vaginal isolation of beta‐haemolytic Streptococcus from bitches with and without neonatal deaths in the litters

The aim of the study was to identify beta‐haemolytic streptococci in the vagina of bitches who had delivered healthy litters and bitches who had delivered litters in which neonatal deaths occurred. Fifty‐one bitches divided into two groups were used. Group 1 (G1) included 28 bitches that had delivered healthy litters and group 2 (G2) included 23 bitches that had delivered puppies who died in the neonatal period. Two vaginal samples were taken, one in proestrus and the other at the end of gestation (EG). Beta‐haemolytic Streptococcus (BS) was isolated from 16 bitches (57%) in G1 and from 21 bitches (91%) in G2. The bacteriological cultures, serological tests (Streptex^®) and PCR assay allowed identification of Streptococcus canis and Streptococcus dysgalactiae in G1 and G2. Ultramicroscopic studies allowed the observation of M Protein and capsules in strains of S. dysgalactiae and S. canis in G1 and G2. The S. canis strains isolated from G2 showed thicker capsules than S. canis strains isolated from G1 (234 ± 24.2 vs 151.23 ± 28.93 nm; p < .001.). No differences were observed in capsule thickness between strains of S. dysgalactiae isolated from G1 and G2 (210 ± 13.54 vs 211.66 ± 19.67 nm; p > .70). All strains of beta‐haemolytic Streptococcus isolated were penicillin sensitive. Penicillin was administered from EG to 5 days post‐partum in 10 G2 females with isolation of BS (G2A). Saline solution was administered in eleven G2 females with isolation of BS (G2B). Ninety per cent of the puppies survived in G2A and 25% survived in G2B. Our results suggest BS is involved in canine neonatal deaths.